Glutamate‐gated N‐methyl‐D‐aspartate (NMDA) channels are the mechanistic link between ammonia and epithelial cell cytotoxicity in Helicobacter pylori infection

Abstract

Background Ammonia (A), a cytotoxin liberated by Helicobacter pylori, is thought to play an important role in cancer development by affecting cell death mechanisms. Our aim was to determine how A is cytotoxic to gastric epithelial cells. Methods Live gastric epithelial cells were used to examine, by microscopy, changes in intracellular Ca2+, mitochondrial membrane potential, and the simultaneous localization of numerous organelle markers in the presence of A with or without specific cellular pathway inhibitors. Biochemical assays were used to quantify cell viability, ATP, protease activation, and the expression of pro-apoptotic proteins. Results A dose-dependently induced Ca2+ influx that was dependent on cAMP. A-induced Ca2+ influx reduced mitochondrial membrane potential and ATP, increased cell death effectors, and activated proteases that overall reduced the viability of gastric epithelial cells. The effects of A could be blocked completely by chelating Ca2+ or by blocking NMDA channels, which are Ca2+ influx channels, with highly specific antagonists. Conclusions Our results are the first to identify NMDA channel activity in epithelial cells, particularly in GI epithelial cells. Our data strongly support that NMDA channels regulate gastric cell viability in the presence of A, rather than other proposed mechanisms including A-induced changes in intracellular pH. Supported by NIH DK-015681.